目的鉴定旋毛虫(Trichinella spiralis,T1)与伪旋毛虫(T.pseudospiralis,T4)肌幼虫的差异蛋白。方法应用SDS-PAGE和双向电泳(two-dimensional gel electrophoresis,2-DE)对T1、T4肌幼虫的可溶性抗原与培养24h的ES抗原的蛋白组分进行分...目的鉴定旋毛虫(Trichinella spiralis,T1)与伪旋毛虫(T.pseudospiralis,T4)肌幼虫的差异蛋白。方法应用SDS-PAGE和双向电泳(two-dimensional gel electrophoresis,2-DE)对T1、T4肌幼虫的可溶性抗原与培养24h的ES抗原的蛋白组分进行分析。结果SDS-PAGE显示,T1肌幼虫可溶性抗原有22条蛋白带(221.62kDa～14.88kDa),其中6条为T1特异性蛋白带(59.72、44.37、23.66、22.36、18.26、16.34kDa);T4可溶性抗原蛋白有18条带(185.28kDa～14.27kDa),其中4条为T4特异性蛋白带(132.60、119.30、35.26、31.02kDa)。T1的ES抗原有10条蛋白带(113.21kDa～14.37kDa),T4的ES抗原有9条蛋白带(104.71kDa～14.51kDa),T1、T4肌幼虫ES抗原的蛋白带均不相同。2-DE显示,T1可溶性抗原有193±12个蛋白点,分子量主要为11kDa～22kDa、25kDa～64kDa及100kDa～144kDa,所对应的等电点(pI)分别为4.7～8.2、4.5～6.5及5～7;T4可溶性抗原有175±9个蛋白点,分子量主要为12kDa～21kDa及25kDa～90kDa,所对应的pI分别为4～9.5与4.5～9.6。T1的ES抗原具有82±6个蛋白点,分子量主要为13kDa～16kDa、18kDa～22kDa及40kDa～55kDa,所对应的pI分别为4～7、3.8～6.2及5～9;T4的ES抗原具有69±5个蛋白点,分子量主要为10kDa～15kDa、17kDa～25kDa及29kDa～55kDa,所对应的pI分别为4.7～6.5、4.6～6及5～7。结论旋毛虫肌幼虫可溶性抗原及ES抗原的蛋白组分与伪旋毛虫的明显不同。展开更多
Objective:To characterize the major allergens of Macrobrachium rosenbergii.(giant freshwater prawn).Methods:Raw and cooked extracts of the giant freshwater prawn were prepared.The IgE reactivity pattern was identified...Objective:To characterize the major allergens of Macrobrachium rosenbergii.(giant freshwater prawn).Methods:Raw and cooked extracts of the giant freshwater prawn were prepared.The IgE reactivity pattern was identified by sodium dodecyl sulfate polyaerylamide gel electrophoresis(SDS-PAGE)and immunoblotting technique with the sera of 20 skin prick test(SPT)positive patients.The major allergen identified was then characterized using the proteomics approach involving a combination of two-dimensional(2-DE)electrophoresis,mass spectrometry and bioinformatics tools.Results:SDS-PAGE of the raw extract showed 23 protein bands(15-250 kDa)but those ranging from 40 to 100 kDa were not found in the cooked extract.From immunoblotting experiments,raw and cooked extracts demonstrated 11 and 5 IgE-binding proteins,respectively,with a molecular mass ranging from 15 to 155 kDa.A heat-resistant 36 kDa protein was identified as the major allergen of both extracts.In addition,a 42 kDa heat-sensitive protein was shown to be a major allergen of the raw extract.The 2-DE gel fractionated the prawn proteins to more than 50 different protein spots.Of these,10 spots showed specific:IgE reactivity with patients'sera.Matrix assisted laser desorption/ionization-lime of flight(MALDI-TOF)analysis led to identification of 2 important allergens,tropomyosin and arginine kinase.Conclusions:It can be concluded that the availability of such allergens would help in component-based diagnosis and therapy of prawn allergies.展开更多
Methicillin-resistant </span><i><span style="font-family:Verdana;">Staphylococcus aureus </span></i><span style="font-family:Verdana;">(MRSA) infection is a glob...Methicillin-resistant </span><i><span style="font-family:Verdana;">Staphylococcus aureus </span></i><span style="font-family:Verdana;">(MRSA) infection is a global health concern that has caused severe health threats over the past decade. Leaves extract of </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">has been proven previously as an anti MRSA agent. Proteomics provide a technique that used to analyze the differential of protein expression profile between untreated and treated MRSA with subinhibitory concentrations of acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span><span style="font-family:Verdana;">leaves. This study aims to determine the optimum parameter for analysis of protein expression profile using two-dimension gels electrophoresis (2-DE) for MRSA protein after treatment with acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllu</span></i></span><i><span style="font-family:Verdana;">m </span></i><span style="font-family:Verdana;">leaves. Comparison of the Protein Expression Profile (PEP) between the untreated and treated MRSA was analyzed using PDQuest software. The optimum condition for MRSA protein treated with acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">leaves to produce the best resolution with greater spot distribution was as follows: 100 μg volume of MRSA protein that loaded after passive rehydration then was run until reaching 25 kVrhs during IEF using 17 cm IPG strip within ranges of pH 4 - 7. Analysis of protein expression from the 2-DE gel map shows that 9 protein spots up-regulated and 41 protein spots were down-regulated with more than 2-fold differences (p < 0.05). This preliminary study on the PEP of MRSA treated with acetone extract of </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">leave may provide an insight into the antimicrobial mechanism, which could lead to the identification of target protein for future novel therapeutic development against MRSA infections.展开更多
该研究采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和双向电泳(2-DE)技术分析比较了鲜冬虫夏草(S1)以及经过22℃阴干(S2)、37℃晒干(S3)和60℃烘干(S4)的冬虫夏草样品中蛋白质成分的变化。Bradford法蛋白含量测定结果显示,4种冬...该研究采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和双向电泳(2-DE)技术分析比较了鲜冬虫夏草(S1)以及经过22℃阴干(S2)、37℃晒干(S3)和60℃烘干(S4)的冬虫夏草样品中蛋白质成分的变化。Bradford法蛋白含量测定结果显示,4种冬虫夏草样品总蛋白质量分数在1. 655~4. 493 mg·g-1,其中鲜冬虫夏草的蛋白含量最高。SDS-PAGE结果显示,冬虫夏草样品蛋白质的相对分子质量主要分布在10~100 k Da,含有的蛋白条带数28~41条,鲜冬虫夏草含有的蛋白条带数量最多。通过PDQuest软件处理冬虫夏草样品2-DE凝胶图谱,结果显示,冬虫夏草蛋白质的等电点主要分布在p I 4. 5~6. 5,相对分子质量主要分布在2个区域10~20 k Da和25~100 k Da,共检测到488~876个蛋白点,鲜冬虫夏草含有的蛋白点数量最多。因此,鲜冬虫夏草含有蛋白成分含量和总类最多,干燥过程会引起冬虫夏草蛋白含量和种类的减少,不同的干燥方法对药材蛋白的影响也不一样,该结果可为冬虫夏草干燥工艺的提升提供参考。展开更多
文摘目的鉴定旋毛虫(Trichinella spiralis,T1)与伪旋毛虫(T.pseudospiralis,T4)肌幼虫的差异蛋白。方法应用SDS-PAGE和双向电泳(two-dimensional gel electrophoresis,2-DE)对T1、T4肌幼虫的可溶性抗原与培养24h的ES抗原的蛋白组分进行分析。结果SDS-PAGE显示,T1肌幼虫可溶性抗原有22条蛋白带(221.62kDa～14.88kDa),其中6条为T1特异性蛋白带(59.72、44.37、23.66、22.36、18.26、16.34kDa);T4可溶性抗原蛋白有18条带(185.28kDa～14.27kDa),其中4条为T4特异性蛋白带(132.60、119.30、35.26、31.02kDa)。T1的ES抗原有10条蛋白带(113.21kDa～14.37kDa),T4的ES抗原有9条蛋白带(104.71kDa～14.51kDa),T1、T4肌幼虫ES抗原的蛋白带均不相同。2-DE显示,T1可溶性抗原有193±12个蛋白点,分子量主要为11kDa～22kDa、25kDa～64kDa及100kDa～144kDa,所对应的等电点(pI)分别为4.7～8.2、4.5～6.5及5～7;T4可溶性抗原有175±9个蛋白点,分子量主要为12kDa～21kDa及25kDa～90kDa,所对应的pI分别为4～9.5与4.5～9.6。T1的ES抗原具有82±6个蛋白点,分子量主要为13kDa～16kDa、18kDa～22kDa及40kDa～55kDa,所对应的pI分别为4～7、3.8～6.2及5～9;T4的ES抗原具有69±5个蛋白点,分子量主要为10kDa～15kDa、17kDa～25kDa及29kDa～55kDa,所对应的pI分别为4.7～6.5、4.6～6及5～7。结论旋毛虫肌幼虫可溶性抗原及ES抗原的蛋白组分与伪旋毛虫的明显不同。
基金Supported by a research grant from UPSI(grant No.UPSI2011-0018-102-01)
文摘Objective:To characterize the major allergens of Macrobrachium rosenbergii.(giant freshwater prawn).Methods:Raw and cooked extracts of the giant freshwater prawn were prepared.The IgE reactivity pattern was identified by sodium dodecyl sulfate polyaerylamide gel electrophoresis(SDS-PAGE)and immunoblotting technique with the sera of 20 skin prick test(SPT)positive patients.The major allergen identified was then characterized using the proteomics approach involving a combination of two-dimensional(2-DE)electrophoresis,mass spectrometry and bioinformatics tools.Results:SDS-PAGE of the raw extract showed 23 protein bands(15-250 kDa)but those ranging from 40 to 100 kDa were not found in the cooked extract.From immunoblotting experiments,raw and cooked extracts demonstrated 11 and 5 IgE-binding proteins,respectively,with a molecular mass ranging from 15 to 155 kDa.A heat-resistant 36 kDa protein was identified as the major allergen of both extracts.In addition,a 42 kDa heat-sensitive protein was shown to be a major allergen of the raw extract.The 2-DE gel fractionated the prawn proteins to more than 50 different protein spots.Of these,10 spots showed specific:IgE reactivity with patients'sera.Matrix assisted laser desorption/ionization-lime of flight(MALDI-TOF)analysis led to identification of 2 important allergens,tropomyosin and arginine kinase.Conclusions:It can be concluded that the availability of such allergens would help in component-based diagnosis and therapy of prawn allergies.
文摘Methicillin-resistant </span><i><span style="font-family:Verdana;">Staphylococcus aureus </span></i><span style="font-family:Verdana;">(MRSA) infection is a global health concern that has caused severe health threats over the past decade. Leaves extract of </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">has been proven previously as an anti MRSA agent. Proteomics provide a technique that used to analyze the differential of protein expression profile between untreated and treated MRSA with subinhibitory concentrations of acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span><span style="font-family:Verdana;">leaves. This study aims to determine the optimum parameter for analysis of protein expression profile using two-dimension gels electrophoresis (2-DE) for MRSA protein after treatment with acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllu</span></i></span><i><span style="font-family:Verdana;">m </span></i><span style="font-family:Verdana;">leaves. Comparison of the Protein Expression Profile (PEP) between the untreated and treated MRSA was analyzed using PDQuest software. The optimum condition for MRSA protein treated with acetone extract from </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">leaves to produce the best resolution with greater spot distribution was as follows: 100 μg volume of MRSA protein that loaded after passive rehydration then was run until reaching 25 kVrhs during IEF using 17 cm IPG strip within ranges of pH 4 - 7. Analysis of protein expression from the 2-DE gel map shows that 9 protein spots up-regulated and 41 protein spots were down-regulated with more than 2-fold differences (p < 0.05). This preliminary study on the PEP of MRSA treated with acetone extract of </span><i><span style="font-family:Verdana;">C</span></i><span style="font-family:Verdana;">.</span><i><span style="font-family:Verdana;"> odontophyllum </span></i><span style="font-family:Verdana;">leave may provide an insight into the antimicrobial mechanism, which could lead to the identification of target protein for future novel therapeutic development against MRSA infections.
文摘该研究采用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和双向电泳(2-DE)技术分析比较了鲜冬虫夏草(S1)以及经过22℃阴干(S2)、37℃晒干(S3)和60℃烘干(S4)的冬虫夏草样品中蛋白质成分的变化。Bradford法蛋白含量测定结果显示,4种冬虫夏草样品总蛋白质量分数在1. 655~4. 493 mg·g-1,其中鲜冬虫夏草的蛋白含量最高。SDS-PAGE结果显示,冬虫夏草样品蛋白质的相对分子质量主要分布在10~100 k Da,含有的蛋白条带数28~41条,鲜冬虫夏草含有的蛋白条带数量最多。通过PDQuest软件处理冬虫夏草样品2-DE凝胶图谱,结果显示,冬虫夏草蛋白质的等电点主要分布在p I 4. 5~6. 5,相对分子质量主要分布在2个区域10~20 k Da和25~100 k Da,共检测到488~876个蛋白点,鲜冬虫夏草含有的蛋白点数量最多。因此,鲜冬虫夏草含有蛋白成分含量和总类最多,干燥过程会引起冬虫夏草蛋白含量和种类的减少,不同的干燥方法对药材蛋白的影响也不一样,该结果可为冬虫夏草干燥工艺的提升提供参考。
